Revised Project Summary/Abstract The overall goal of the proposed research is to identify epigenetic changes in biological pathways associated with protection from Type 2 Diabetes (T2D) in Yup?ik Alaska Native people consuming a traditional diet. The Yup?ik traditional dietary pattern is characterized by high protein, high fat, and low carbohydrate intakes, as well as n-3 polyunsaturated fatty acid levels that are up to 20 times higher than in the general U.S. population. High levels of traditional food intake in Yup?ik people are positively associated with adiponectin and inversely associated with c-reactive protein, triglycerides, and the leptin:adiponectin ratio, suggestive of increased insulin sensitivity. However, the diet of Yup?ik people is transitioning to increased consumption of highly processed market foods, and the prevalence of overweight and obesity is beginning to mirror that of the general U.S. population. Even so, the prevalence of metabolic syndrome (15%) and T2D (3%) among Yup?ik people remains lower than in the general U.S. population. Adherence to a Yup?ik traditional dietary pattern is likely to be protective in Yup?ik people, although the mechanisms of protection are not well understood. We will use the nitrogen stable isotope ratio (?15N) as a validated, objective measure of the traditional dietary pattern to explore these mechanisms. Although considerable advances have been made in the identification of genetic factors contributing to metabolic diseases, less than 10% of the heritability of T2D phenotypes is explained by known genetic variation. Epigenetic mechanisms have recently been proposed as a link between genetic risk and environmental exposures such as diet, thereby providing a mechanism for the modification of genetic predisposition and insight into 90% of the heritability that is not explained. The proposed research will fine map epigenetic biomarkers in the 20 most promising candidate genes identified using Illumina MethylationEPIC microarrays on stored peripheral blood DNA samples from 569 Yup?ik people at the highest and lowest deciles of traditional dietary intake (DK104347) using targeted bisulfite sequencing to reveal detailed DNA methylation patterns not apparent in the array data, and to identify genetic variants that influence methylation at these genes. These findings will be replicated in a Greenlandic Inuit cohort to determine the generalizability of the findings from Yup?ik people. The downstream functional consequences of methylation patterns in the candidate genes will be assessed using gene expression assays in 192 peripheral blood RNA samples from individuals at the extremes of the distribution of our dietary measure, ?15N. By identifying epigenomic biomarkers associated both with adherence to a traditional diet and with insulin sensitivity, the proposed research will test the hypothesis that a traditional Yup?ik diet modifies the downstream transcription of key genes that contribute to metabolic health of Yup?ik people.